1. Field of the Invention
The present invention relates to a novel maltohexaose and maltoheptaose-forming amylase, and preparation and uses thereof, more particularly, to a maltohexaose and maltoheptaose-forming amylase which mainly forms maltohexaose and maltoheptaose from starch (designated as "AMYLASE" hereinafter) and preparation thereof, as well as microorganisms which produce such an amylase, saccharide compositions containing maltohexaose and/or maltoheptaose produced therewith or those containing maltohexaitol and/or maltoheptaitol obtainable by hydrogenating the maltohexaose and maltoheptaose, and compositions containing these saccharides.
2. Description of the Prior Art
Methods for specifically producing maltooligosaccharides such as maltose and maltotetraose by allowing specific amylases to act on starch have been employed on an industrial scale, and widely used in processes of compositions such as foods and pharmaceuticals. Demands for saccharides containing a relatively-large quantity of maltooligosaccharides such as maltohexaose and maltoheptaose have increased because, among maltooligosaccharides, those with a relatively-low molecular weight are considerably-low in sweetness, readily digestible and absorbable.
Enzymes or amylases, derived from microorganisms, which produce a relatively-large amount of maltohexaose when act on starch are known, but no amylase which forms maltooligosaccharides higher than maltoheptaose has been known.
Amylases which mainly produce maltohexaose from starch can be roughly classified into two groups based on their actions. The one is an exo-type maltohexaose-forming amylase which hydrolyzes amylaceous substances so as to remove successive maltohexaose units from the non-reducing chain ends, namely, maltohexaohydrolase (EC 3.2.1.98), and the other is an endo-type maltohexaose-forming amylase, namely, endo-type .alpha.-amylase (EC 3.2.1.1) which acts on internal amylaceous saccharide chains to produce maltohexaose in a relatively-large quantity.
Kainuma et al. reported in FEBS Letters, Vol.26, pp.281-285 (1972) a microorganism of the species Aerobacter aerogenes which intracellularly forms an exo-type maltohexaose-forming amylase which is characteristic of its relatively-low optimum temperature and thermal stability, but it is insufficient in heat tolerance for an industrial use.
J. F. Kennedy et al. reported in Starch, Vol.31, pp.235-241 (1979) a microorganism of the species Bacillus subtilis forms an endo-type maltohexaose-forming amylase, and Takasaki reported in Agricultural and Biological Chemistry, Vol.46, pp.1539-1547 (1982) a microorganisms of Bacillus circulans G-6 forms such an amylase. Taniguchi reported in Denpun Kagaku (Starch Science), Vol.29, pp.107-116 (1982) a microorganism of the species Bacillus circulans F-2 forms such an amylase, and Hayashi et al. reported in Agricultural and Biological Chemistry, Vol.52, pp.443-448 (1988) a microorganism of the species Bacillus sp. H-167 forms such an amylase. It has been known that, among these amylases produced from such microorganisms, those from Bacillus circulans G-6, Bacillus circulans F-2 and Bacillus sp. H-167 form maltohexaose from starch in the maximum yield of about 25-30 w/w %, on a dry solid basis (d.s.b.), (the wording "w/w %" is designated as "%" hereinafter, if not specified otherwise), but they do not form maltoheptaose, and further they hydrolyze maltohexaose into maltose and maltotetraose as their enzymatic reactions proceed.
Now referring to amylases derived from microorganisms of the species Bacillus subtilis, they only form about 25% maltohexaose from starch and do not hydrolyze maltohexaose. Such amylases are not suitable for an industrial-scale production of high maltohexaose content saccharides because they do not form maltohexaose in a satisfactorily-high yield while forming a relatively-large quantity of lower molecular oligosaccharides under starch saccharification conditions, and because they could not be readily obtained in quantity from microorganisms.
In Agricultural and Biological Chemistry, Vol.42, pp.259-267 (1978) it was reported that a purified .alpha.-amylase, prepared from malts, mainly forms maltohexaose and maltoheptaose during the early stage of the enzymatic reaction on starch. It is also reported that the formed maltohexaose and maltoheptaose are decomposed almost completely into lower molecular oligosaccharides such as maltose and maltotetraose as the enzymatic reaction proceeds, and, therefore, it scarcely produces saccharified products, mainly containing maltohexaose and maltoheptaose, from starch.